Tuberculosis, one of the great scourges of humankind, continues to be a serious and significant health problem in the world today. The World Health Organization estimates that one-third of the world's population is infected with Mycobacterium tuberculosis, the causative agent of TB. Approximately 8 million new cases of TB are diagnosed annually worldwide. Of those individuals with active disease, nearly 2 million die each year. Small wonder, then that in 1993, WHO declared tuberculosis to be a global health emergency, as it remains today. During the past 12-14 years, significant progress has been made in the development of genetic tools to facilitate analysis of the genes and gene products of M. tuberculosis. The nucleotide sequences of the entire genomes of two strains of M. tuberculosis have been determined. The techniques of DNA microarray analysis and two-dimensional gel electrophoresis analysis have provided valuable information about global gene expression and protein production by M. tuberculosis grown in defined environments. We developed a method to identify M. tuberculosis genes that are expressed when the mycobacteria are growing in their favored ecological niche, the human macrophage. Although much has been learned, fundamental aspects of M. tuberculosis pathogenesis remain unknown. Employing a number of currently available genetic, genomic, proteomic, and molecular biological tools, we propose to (1) characterize the regulons of the TrcRS, Rv1626, and PrrAB regulatory systems, (2) determine the contribution of specific gene products to M. tuberculosis pathogenesis by comparisons between wild-type M. tuberculosis H37Rv and strains of H37Rv with mutations in specific genes expressed during growth in human macrophages and (3) compare gene expression by M. tuberculosis growing in human peripheral blood monocyte-derived macrophages and in infected mouse lungs by cDNA microarray and quantitative RT-PCR (QRT-PCR) analyses.